Human and mouse embryonal carcinoma cells and early embryo cells display a unique phenotype: they do not express certain viral and cellular genes that are expressed in differentiated successors. Therefore, they provide a system to study interesting aspects of the control of gene expression during very early embryonic development. We have found evidence that primary control of gene expression in the EC cell occurs by mechanisms other than de novo methylation. Although de novo methylation occurs, it may be a consequence of nonexpression. We will examine the effects of introducing CAD or other genes into embryonal carcinoma cells to determine whether they are expressed at any time or all times after introduction to the cells. The sequences will be transformed into the cells by calcium phosphate co-precipitation or protoplast fusion. Does the expression of introduced sequences change with integration into the genome of the mouse cell? Does the expression correlate with methylation patterns of the sequences? We will determine whether embryonal carcinoma cells produce progeny virus after infection with MoMLV variants that we have constructed which contain mutant polyoma virus-enhancer region sequences in the LTRs. If progeny virus is made, then it will be clear that the block to proviral gene expression in embryonal carcinoma cells is due to the inability of the EC cell to utilize the endogenous or wt MoMLV transcriptional enhancer or activator sequences. Based on the results, we will construct a retrovirus vector containing the proper regulatory sequences that will allow expression of cloned genes introduced into EC or preimplantation embryo cells by infection with this vector. (M)